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AlCapone2k
Beginner


Registered: 10/06/16
Posts: 842
Loc: Germany
Last seen: 6 years, 4 months
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NoobLog: Print -> Agar -> Rye Spawn -> CVG -> Mono
#23765984 - 10/24/16 05:38 AM (7 years, 3 months ago) |
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Hey guys,
since I have asked a lot of questions in the Question and Answer Threads, I thought to start with a small log where I can show you my results so far and if there's a question, I can ask it directly here. Maybe some of the great growers here can help me along 
My Plan:Spore Print --> Agar --> Agar Transfers and Isolation until good ryzomorphic mycelium --> Agar wedge inoculation of rye jar --> Colonization --> Bulk --> Monotub.
First of all: I'm from Germany, my english is quite good, but sometimes I don't understand urban language, so don't be mad when I ask you to repeat some sentences in other words if I didn't understand them 
Ok, let's start.
Two monts ago I bought a Grow Kit, was my first one, before I never got in contact with psychedelic fungi. The growkit was just fine, gave me a great first flush of 10g cracker dried. The second flush was not that good, gave just a couple of very big shrooms but I took the chance to make a sporeprint.
SAB-Tek used: Spitball Jedi (https://www.shroomery.org/forums/showflat.php/Number/20048771#20048771)
Pour Agar Tek: Bhodisatta (https://www.shroomery.org/forums/showflat.php/Number/21922023)
1. Sporeprint making (10.10.2016) I made a sporeprint with the help of bodhisatta in my SAB. Everything worked well so I had two good sporeprints. I skipped to make a spore syringe cause I directly started with agar.
2. Pour agar (15.10.2016) I made a LME-Agar and poured it into my petri dishes. Most plates were poured more than ten days ago and since now there is no contamination on them, so it seems, that I worked quite sterile while pouring the agar plates.
3. Inoculation of the agar plates (17.10.2016) I used sterile cotton swabs, wettened them a bit on the agar plate and then I got it on the spore print to grab some spores.
I've inoculated 4 agar plates.
2 with striking in a z-pattern and 2 with circle movement in the middle of the plate. I wrapped two with saran wrap and two with parafilm.
These are the results after one week (7 days) now:
BETTER PICTURES ARE SOME POSTES BELOW
Picture 1: Striking + Saran

Picture 2: Striking + Saran

Picture 3: Circle Movement + Parafilm

Picture 4: Circle Movement + Parafilm

After one week after inoculation they show as expected a good mycelium grow. Till now I can't see any contamination, but to be honest, I actually don't know whether I could recognize one if I see it.
So my first questions:
1. Can you see any contamination? Does everything look good? Is it fine until now? Too much or to less mycelium grow after a week? Is it normal that at the point where I put the spores onto there is dense white mycelium and at the edges its getting less dense and less white?
2. Is it a problem that there seem to be some spores not germinated (black dots in the mycelium)?
3. Do you think I should start with "cleaning"-transfer to other plates or just wait a bit? Shall I go with one plate, transfer to other agar and meanwhile just let it (the one i transferred from) colonize further to go for an isolation and rye inoculation?
Or shall I only start isolation after some cleaning-transfers?
4. If I want to transfer for cleaning, how big shall the wedge to cut out be? As small as possible? From the very edge of a myelium dot or better from the middle where it's dense and white?
Thanks in advanced for any help!
Regards.
-------------------- Since I am from Germany, my english is not perfect but I try my best    My small Agar to Oyster Open Air Grow Trek
Edited by AlCapone2k (10/24/16 07:48 AM)
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AgarStudent
Noob


Registered: 01/25/15
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Re: NoobLog: Print -> Agar -> Rye Spawn -> CVG -> Mono [Re: AlCapone2k]
#23766002 - 10/24/16 06:05 AM (7 years, 3 months ago) |
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Hmm if you are serious about the hobby, I would recommend a laminar flow hood to avoid contaminations. You can build one yourself for not that much.
I don't know how you can avoid contamination without one to be honest, but some people seem to do without them? Your plates look okay, based on the resolution of the images you have provided.
I would wait until you have a bit more mycelial growth before you do any transfers. Maybe another few days to a week.
If you have good growth, somewhere on the edge would be best for a transfer. I think this is the fastest growing but of mycelium.
-------------------- Why reinvent the wheel when someone here has already done the science?
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PortabellaFella 1
Enthusiastic



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Re: NoobLog: Print -> Agar -> Rye Spawn -> CVG -> Mono [Re: AlCapone2k]
#23766005 - 10/24/16 06:09 AM (7 years, 3 months ago) |
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1. I can't really see much. The best way to take a pic of a plate is with a light shined at the side of the plate, they are bit blurry yes it's normal to be dense at the center and decrease outward.
2. You shouldn't be able to see black spores with the naked eye unless you literally didn't shake the syringe and a giant mass came out
3. You can transfer to more agar but I'd suggest taking a few transfers from a single individual plate think of a pyramid where the top is your original plate, everything below the "point" are transfers. You could let the original plate colonize if you don't need the plate and maybe it will pin. If it does transfer or "clone" the pin and then try to isolate. It's rare and highly unlikely that you will get an isolate directly from multispore and would take a shit ton of transfers to get one.
4. You transfer from a clean edge of the mycelium growth, with multispore your not going to see a lot if any sectoring so you really won't know what you're transferring. Your transfer wedge doesn't need to be any specific size right now, just make it big enough to get a good hold on the tip of your heat flamed then cooled scalpel.
-------------------- I would like to acquire anything I don’t have
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thebug76
2 years in.



Registered: 05/31/15
Posts: 557
Loc: North of the equater and ...
Last seen: 3 months, 26 days
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Re: NoobLog: Print -> Agar -> Rye Spawn -> CVG -> Mono [Re: PortabellaFella 1]
#23766027 - 10/24/16 06:28 AM (7 years, 3 months ago) |
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I'm seeing what looks like some mold in all the plates (dead center, 7:30ish, and 1:00 in pic 1; 10:00 in pic 2; 2 spots just above dead center in pic 3; and 8 or 9 spots around center in pic 4), but like the other guys said, the pics aren't great. I also see what looks like clean cube mycelium. Get some better pics and the guys on here will be able to help a lot more. Plus that kind of growth after only 1 week is either some damn good spores, or isn't cube mycelium. Mine usually take longer than a week to even germinate, then I'm looking at another 2-3 days to get that kind of growth.
-------------------- Bug
Edited by thebug76 (10/24/16 06:40 AM)
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thebug76
2 years in.



Registered: 05/31/15
Posts: 557
Loc: North of the equater and ...
Last seen: 3 months, 26 days
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Re: NoobLog: Print -> Agar -> Rye Spawn -> CVG -> Mono [Re: PortabellaFella 1]
#23766043 - 10/24/16 06:37 AM (7 years, 3 months ago) |
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Quote:
PortabellaFella 1 said:
2. You shouldn't be able to see black spores with the naked eye unless you literally didn't shake the syringe and a giant mass came out
He said he skipped the syringe and went cottonswab to print to agar. Thought I'd point that out, it could explain the spore clumps, but it kind of looks like some mold growth. I pointed it out in my previous reply, I totally missed you suggesting OP didn't shake the syringe or I would be pointed that out then also.
-------------------- Bug
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AlCapone2k
Beginner


Registered: 10/06/16
Posts: 842
Loc: Germany
Last seen: 6 years, 4 months
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Re: NoobLog: Print -> Agar -> Rye Spawn -> CVG -> Mono [Re: AlCapone2k]
#23766056 - 10/24/16 06:49 AM (7 years, 3 months ago) |
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Thank you very much for your answers.
Here are some better Pictures I hope:
Picture 1:
Picture 2:

Picture 3:

Picture 4:

I hope they are better now
To your postings:
Quote:
Hmm if you are serious about the hobby, I would recommend a laminar flow hood to avoid contaminations. You can build one yourself for not that much.
Maybe in some weeks I will build one, but at the moment it must work with "only" a SAB.
Quote:
2. You shouldn't be able to see black spores with the naked eye unless you literally didn't shake the syringe and a giant mass came out
I made it with a spore print and a cotton swab, the cotton swab was black when touching the agar plate, so thats the reason for the black dots. Some germinated and disappeared but some stay.
Quote:
It's rare and highly unlikely that you will get an isolate directly from multispore and would take a shit ton of transfers to get one.
So the first step should be cleaning-transfer and isolation is something I can think about after first grow and cloning?
Quote:
I'm seeing what looks like some mold in all the plates. Plus that kind of growth after only 1 week is either some damn good spores, or isn't cube mycelium. Mine usually take longer than a week to even germinate, then I'm looking at another 2-3 days to get that kind of growth.
Really? Also on the new pictures? Damn
-------------------- Since I am from Germany, my english is not perfect but I try my best    My small Agar to Oyster Open Air Grow Trek
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PortabellaFella 1
Enthusiastic



Registered: 08/08/16
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Re: NoobLog: Print -> Agar -> Rye Spawn -> CVG -> Mono [Re: thebug76]
#23766064 - 10/24/16 06:59 AM (7 years, 3 months ago) |
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It's early, I skipped over the cotton swab inoculation my bad. That's myc. It's gonna be a little difficult to transfer from a leading edge since there are so many colonies with the spores swabbed so thick. Yes, clean, transfer, grow , clone then isolate. The new pics are much better.
-------------------- I would like to acquire anything I don’t have
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thebug76
2 years in.



Registered: 05/31/15
Posts: 557
Loc: North of the equater and ...
Last seen: 3 months, 26 days
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Re: NoobLog: Print -> Agar -> Rye Spawn -> CVG -> Mono [Re: PortabellaFella 1]
#23766081 - 10/24/16 07:12 AM (7 years, 3 months ago) |
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The new pics look better. I think some of the spots I was seeing is either clumps or dark spots on the lids of your plates. On the new pics there's a couple of spots in pic 2 that are either heavy spore clumps or are about to turn green. But even if it does, you still have good stuff to work from.
-------------------- Bug
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AlCapone2k
Beginner


Registered: 10/06/16
Posts: 842
Loc: Germany
Last seen: 6 years, 4 months
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Re: NoobLog: Print -> Agar -> Rye Spawn -> CVG -> Mono [Re: thebug76]
#23766127 - 10/24/16 07:52 AM (7 years, 3 months ago) |
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Quote:
PortabellaFella 1 said: It's early, I skipped over the cotton swab inoculation my bad. That's myc. It's gonna be a little difficult to transfer from a leading edge since there are so many colonies with the spores swabbed so thick. Yes, clean, transfer, grow , clone then isolate. The new pics are much better.
hehe I just forgot that it's early in the morning in the US, here it's 4 pm
But the plate on picture 4 - can I transfer from the leading edge there?
How shall I transfer from the other pleates? Directly a wedge with a whole "myelium dot"?
Shall I transfer as soon as possible or wait a bit? Which points do you recommend for transfer?
Quote:
thebug76 said: The new pics look better. I think some of the spots I was seeing is either clumps or dark spots on the lids of your plates. On the new pics there's a couple of spots in pic 2 that are either heavy spore clumps or are about to turn green. But even if it does, you still have good stuff to work from.
Thank you very much, I was really afraid that it's mold, cause with my eyes I can't see anything strange.
Looking at plate of picture two I cant see anything turnin green. Maybe it's the picture, but I will observe it.
"To work with" means cleaning-tranfers, right? Shall I start asap or wait a bit?
Which points do you recommend to transfer?
-------------------- Since I am from Germany, my english is not perfect but I try my best    My small Agar to Oyster Open Air Grow Trek
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PortabellaFella 1
Enthusiastic



Registered: 08/08/16
Posts: 654
Last seen: 2 years, 9 months
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Re: NoobLog: Print -> Agar -> Rye Spawn -> CVG -> Mono [Re: AlCapone2k]
#23766143 - 10/24/16 08:03 AM (7 years, 3 months ago) |
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Picture 4 I would cut a wedge from the top colony about 1 o'clock. Just take a wedge from the leading edge inward toward the center but away from the fluffy white mass. You could do it now or wait to see if any more aggressive myc forms.
-------------------- I would like to acquire anything I don’t have
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thebug76
2 years in.



Registered: 05/31/15
Posts: 557
Loc: North of the equater and ...
Last seen: 3 months, 26 days
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Re: NoobLog: Print -> Agar -> Rye Spawn -> CVG -> Mono [Re: PortabellaFella 1]
#23766157 - 10/24/16 08:15 AM (7 years, 3 months ago) |
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Right around the 12:00 position in pic 2 looks really thick and kind of dark, that's where I said possibly mold. Could be lighting tho. But in any case you have good rhizo growth near the bottom and just above center. There's several places you could get good transfers from, just get from the leading edge. I don't even really do wedges when I'm making plate to plate transfers, only a really small piece about the size of a grain of rice. Less is more when going agar->agar.
I would go ahead and do it. If you wait much longer, that plate is going to be all grown together and overrun.
-------------------- Bug
Edited by thebug76 (10/24/16 08:17 AM)
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AlCapone2k
Beginner


Registered: 10/06/16
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Loc: Germany
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Re: NoobLog: Print -> Agar -> Rye Spawn -> CVG -> Mono [Re: thebug76]
#23766319 - 10/24/16 09:57 AM (7 years, 3 months ago) |
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Thank you for your answers.
Shall I also transfer from plate on picture 4? The mycelium is not that big on that but at least some kind of rhizomorphic
-------------------- Since I am from Germany, my english is not perfect but I try my best    My small Agar to Oyster Open Air Grow Trek
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thebug76
2 years in.



Registered: 05/31/15
Posts: 557
Loc: North of the equater and ...
Last seen: 3 months, 26 days
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Re: NoobLog: Print -> Agar -> Rye Spawn -> CVG -> Mono [Re: AlCapone2k]
#23766382 - 10/24/16 10:35 AM (7 years, 3 months ago) |
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I would go with portabellafella1's advice on pic 4. I've read a lot of his stuff on here and honestly, he's probably more qualified than I am.
-------------------- Bug
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AlCapone2k
Beginner


Registered: 10/06/16
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Re: NoobLog: Print -> Agar -> Rye Spawn -> CVG -> Mono [Re: thebug76]
#23766460 - 10/24/16 11:12 AM (7 years, 3 months ago) |
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Quote:
thebug76 said: I would go with portabellafella1's advice on pic 4. I've read a lot of his stuff on here and honestly, he's probably more qualified than I am.
Ok, fine, and what do you think of the other plates, also transfer?
I will update a picture where I mark the transfer points right now
-------------------- Since I am from Germany, my english is not perfect but I try my best    My small Agar to Oyster Open Air Grow Trek
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thebug76
2 years in.



Registered: 05/31/15
Posts: 557
Loc: North of the equater and ...
Last seen: 3 months, 26 days
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Re: NoobLog: Print -> Agar -> Rye Spawn -> CVG -> Mono [Re: AlCapone2k]
#23766488 - 10/24/16 11:23 AM (7 years, 3 months ago) |
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Anywhere you have clean looking rhizomorphic growth, you can transfer from. I wouldn't get too transfer happy tho until I had a few plates gaurenteed clean. And even then, it doesn't take much. It takes a bunch of plates when you're isolating. Plates grow fairly quick once you get them going, so it's not like tou need buttloads sitting around. Making sure you have a clean culture can generally be done in just a few transfer.
-------------------- Bug
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Greg
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Re: NoobLog: Print -> Agar -> Rye Spawn -> CVG -> Mono [Re: AlCapone2k]
#23766493 - 10/24/16 11:27 AM (7 years, 3 months ago) |
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All of those plates look good to me. I feel like taking transfers from more than one plate is redundant at this stage because it's all random as hell MS anyway. Just focus on getting clean healthy growth.
Generally what I do is I make 2 germination plates, take 2-4 transfers from the best looking one, take 2-4 transfers from the best looking one of those, etc. I always keep the old plates until I am certain I like the new transfers so I can go back a step if needed.
Don't worry if you can't get rhizo growth at first (or at all), tomentose growth can be healthy too. My burma took ~5 transfers to show rhizo traits and it still reverts back to tomentose when I transfer to new plates sometimes. On the other hand, my KSSS showed rhizo growth on the first transfer and my APE showed it on the germination plate. Point is, it doesn't necessarily matter if it's rhizo, it only matters if it's healthy.
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PortabellaFella 1
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Re: NoobLog: Print -> Agar -> Rye Spawn -> CVG -> Mono [Re: Greg]
#23766503 - 10/24/16 11:33 AM (7 years, 3 months ago) |
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-------------------- I would like to acquire anything I don’t have
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AlCapone2k
Beginner


Registered: 10/06/16
Posts: 842
Loc: Germany
Last seen: 6 years, 4 months
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Re: NoobLog: Print -> Agar -> Rye Spawn -> CVG -> Mono [Re: Greg]
#23766529 - 10/24/16 11:43 AM (7 years, 3 months ago) |
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Quote:
Greg said: All of those plates look good to me. I feel like taking transfers from more than one plate is redundant at this stage because it's all random as hell MS anyway. Just focus on getting clean healthy growth.
Generally what I do is I make 2 germination plates, take 2-4 transfers from the best looking one, take 2-4 transfers from the best looking one of those, etc. I always keep the old plates until I am certain I like the new transfers so I can go back a step if needed.
Don't worry if you can't get rhizo growth at first (or at all), tomentose growth can be healthy too.
Thank you for your answer (and of course thanks to all the others).
So you suggest to only transfer from one dish is better than from all the dishes, right? Which one would you chose? Also plate on Picture 4?
And you cut more than one wedge of ONE plate and transfer to different new plates, did I understand correctly? So you make e.g. 3 plates out of one? Isn't it a contamination risk if I open the plate that long to take for example 3 wedges out of it?
Here I have a picture of plate 4. Which one of the triangles would you choose for transferring? Just to get an idea how near to the "center" of the growth I can go.
-------------------- Since I am from Germany, my english is not perfect but I try my best    My small Agar to Oyster Open Air Grow Trek
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thebug76
2 years in.



Registered: 05/31/15
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Re: NoobLog: Print -> Agar -> Rye Spawn -> CVG -> Mono [Re: AlCapone2k]
#23766542 - 10/24/16 11:49 AM (7 years, 3 months ago) |
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All three look clean. Me personally, I would go with red. Either would be sufficient, but newer growth tends to be more vigorous. Then I would get one from around the bottom and from the upper left as well.
-------------------- Bug
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blackout


Registered: 07/16/00
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Re: NoobLog: Print -> Agar -> Rye Spawn -> CVG -> Mono [Re: AgarStudent]
#23766550 - 10/24/16 11:51 AM (7 years, 3 months ago) |
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Quote:
AlCapone2k said: Two monts ago I bought a Grow Kit, was my first one, before I never got in contact with psychedelic fungi. The growkit was just fine, gave me a great first flush of 10g cracker dried. The second flush was not that good, gave just a couple of very big shrooms but I took the chance to make a sporeprint.
I would have taken a clone of that growkit. It is a known fruiting strain which was presumably carefully selected.
Seems you were streaking the plates and having growth in many places. I would only dip the swab in the very centre, this way if any spots other than the centre show growth it would be unwanted growth and you know your SAB technique is not great, or that your plates were not sterile to start with. It also means it will grow outwards from the centre so its is easier to select a clean leading edge. Having many dots of growth means they will grow into each other. Instead of dipping swabs I have picked a little bit off with a sterile tweezers and placed it in the middle of a plate, I did this as my plates were quite moist and I was worried about it absorbing nutrients, though others say they have no issue.
Quote:
AgarStudent said: Hmm if you are serious about the hobby, I would recommend a laminar flow hood to avoid contaminations. You can build one yourself for not that much.
I don't know how you can avoid contamination without one to be honest, but some people seem to do without them?
Surely you have heard of SABs? I have been simply using a long clear plastic bag lately, which is sprayed with water..
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